Department of Life Science, Pohang University of Science and Technology, Pohang 37673, Korea.
Int J Mol Sci. 2020 May 8;21(9):3332. doi: 10.3390/ijms21093332.
Serial crystallography (SX) provides an opportunity to observe the molecular dynamics of macromolecular structures at room temperature via pump-probe studies. The delivery of crystals embedded in a viscous medium via an injector or syringe is widely performed in synchrotrons or X-ray free-electron laser facilities with low repetition rates. Various viscous media have been developed; however, there are cases in which the delivery material undesirably interacts chemically or biologically with specific protein samples, or changes the stability of the injection stream, depending on the crystallization solution. Therefore, continued discovery and characterization of new delivery media is necessary for expanding future SX applications. Here, the preparation and characterization of new polysaccharide (wheat starch (WS) and alginate)-based sample delivery media are introduced for SX. Crystals embedded in a WS or alginate injection medium showed a stable injection stream at a flow rate of < 200 nL/min and low-level X-ray background scattering similar to other hydrogels. Using these media, serial millisecond crystallography (SMX) was performed, and the room temperature crystal structures of glucose isomerase and lysozyme were determined at 1.9-2.0 Å resolutions. WS and alginate will allow an expanded application of sample delivery media in SX experiments.
连续结晶学 (SX) 通过泵浦探测研究提供了在室温下观察大分子结构分子动力学的机会。通过注射器或注射器将嵌入在粘性介质中的晶体输送到同步加速器或 X 射线自由电子激光设施中，这些设施的重复率较低。已经开发了各种粘性介质；然而，在某些情况下，输送材料会与特定的蛋白质样品发生不理想的化学或生物相互作用，或者根据结晶溶液改变注射流的稳定性。因此，需要不断发现和表征新的输送介质，以扩展未来的 SX 应用。在这里，介绍了用于 SX 的新型多糖（小麦淀粉（WS）和藻酸盐）基样品输送介质的制备和特性。嵌入 WS 或藻酸盐注射介质中的晶体在<200nL/min 的流速下显示出稳定的注射流，并且与其他水凝胶相似的低水平 X 射线背景散射。使用这些介质进行了连续毫秒结晶学 (SMX)，并在 1.9-2.0Å分辨率下确定了葡萄糖异构酶和溶菌酶的室温晶体结构。WS 和藻酸盐将允许在 SX 实验中扩展样品输送介质的应用。
